New growths produced by the expansion of local tissue cells in response to many tumorigenic triggers are referred to as “tumors”. It is primarily a mass protrusion that occupies space, also known as a neoplasm [1,2]. In terms of morbidity and mortality, cancer continues to be a major global public health issue, coming in second place only behind cardiovascular disease [3,4]. A report on worldwide cancer statistics published in 2020 by the International Agency for Research on Cancer indicated that, in this century, cancer might surpass cardiovascular disease as the major cause of premature death in the majority of countries. By 2040, there are projected to be 28.4 million new cases of cancer worldwide, an increase of 47% from 2020 [5,6]. The pathogenic mechanisms of cancer include maintaining proliferative signals, eluding growth inhibitors, avoiding cell death, establishing replicative immortality, initiating angiogenesis, and triggering invasion and metastasis [7]. Over the last 50 years, a lot of strategies have been employed to slow the spread of cancer, namely surgery, radiotherapy, and systemic therapy [8,9]. These treatments, however, have numerous limitations and side effects, such as a high incidence of drug resistance and multidrug resistance, low efficacy of some targeted therapies, and severe adverse responses, whether administered alone or in combination [10,11]. Thus, developing novel anti-tumor medications and minimizing side effects have consistently been major areas of research in the anti-tumor field.

Due to their multi-channel and multi-target characteristics, anti-tumor compounds derived from natural products are typically used to treat advanced cancer and relieve early cancer symptoms. Herbs used in traditional medicine are the source of most natural products [12–14]. Over the past few decades, natural products have been a significant source for the development of new anti-tumor drugs [11]. Currently, more than 100 natural compounds are clinically used to treat cancers [15,16]. For example, studies have demonstrated that the active ingredient in Curcuma longa, curcumin, exerts anti-tumor effects by boosting apoptosis, inhibiting cell proliferation, obstructing tumor angiogenesis and metastasis, and inducing autophagy [17,18]. For example, paclitaxel exerts its anti-breast cancer effects by blocking mitosis (affecting B-cell lymphoma 2 (Bcl-2) phosphorylation), controlling microtubule polymerization, affecting calcium signaling, and regulating microRNA expression profiles [19]. Furthermore, significant anti-cancer efficacy is exhibited by artemisinin, which can generate reactive oxygen species in cancer cells, induce cell cycle arrest and autophagy, block cancer cell invasion and migration, and accelerate cancer cell apoptosis [20].

In recent years, a lot of literature has been reported about the classical anti-tumor signaling pathways of natural products, which are found to have the advantages of multiple targets and pathways, providing a sufficient theoretical basis for natural products as more promising antitumor drugs. For instance, the phosphoinositide 3-kinase (PI3K) signaling pathway is a crucial signaling pathway for normal cell physiological metabolism. By inhibiting this signaling pathway, tumor growth and metastasis can be prevented. Activation of PI3K can also assist in the phosphorylation of Akt and, ultimately, together play a crucial role in tumor progression [21]. For instance, Yan et al. [22] found that baicalein induced apoptosis and autophagy in breast cancer cells by inhibiting PI3K/Akt signaling pathway. In addition to the PI3K signaling pathway, the signal transducer and activator of the transcription 3 (STAT3) signaling pathway is one of the key targets for cancer therapy, which is a significant intracellular signal transduction protein and transcription factor directly related to the growth of tumors [23]. Luo et al. [24] found that Bavachin induces iron death in osteosarcoma cells via inhibition of STAT3 activity. Additionally, mitogen-activated protein kinase (MAPK) is a crucial intracellular signal transduction system that controls a variety of processes in cells, including cell growth, proliferation, differentiation, apoptosis, adhesion, and migration. These processes have an impact on drug resistance, invasion, and metastasis, as well as tumorigenesis, making it one of the potential targets of anti-tumor medications [25].

With more than 700 species recognized, the genus Selaginella of the family Selaginellaceae is extensively distributed around the world; especially, it is primarily found in tropical and subtropical areas. These species have been reported to possess many bioactivities that include anticancer, anti-inflammatory, antibacterial, antiviral, antioxidant, anti-aging, hypoglycemic, and other activities [26,27]. However, the current pharmacological studies of Selaginella plants are mainly focused on anti-tumor effects. For example, Ethyl acetate fractions from Selaginella doederleinii exhibited cytotoxicity effects for A549 cell lines, 7721 cell lines, Hela cell lines and Eca-109 cell lines, among which it has the strongest inhibitory effect on Hela cell lines with IC50 value of 37.53 μg/mL [28]. Li et al. [29] discovered that ethyl acetate extract from Selaginella doederleinii induced autophagic death and apoptosis in colorectal cancer cells through the PI3K-Akt-mTOR and AMPKα signaling pathways. Lei J et al. [30] found that the biflavonoid extracts from S. moellendorffii exhibited a noticeable negative effect on the growth rate of HCT-116 cell lines and HeLa cell lines in the range of 0 μg/mL to 500 μg/mL. Qin et al. [31] used proline-lactic acid to prepare biflavonoid extracts, which showed significant inhibitory activity against tumor A549 cells, sw1990 cells and HepG2 cells. Therefore, we comprehensively discussed biflavonoids from Selaginella, which are the main active ingredient that exerts anti-tumor effects and devoted ourselves to elucidating its anti-tumor mechanism of action and its signaling pathway in the review. As shown in Figure 1.

In the last two decades, scientists have discovered that Selaginella plants have a wide range of active components, mostly classified as alkaloids, selaginellins and phenolic acids. Selaginellin components have been found to have potential anti-tumor activity due to their unique triple-bond structure. For example, Zhang et al. [32] isolated five selaginellin derivatives from Selaginella tamariscina, and two of the new selaginellins were structurally characterized by spectroscopic analysis. Cytotoxic activity assessment showed that the two new selaginellins exhibited moderate toxicity against human cancer cell lines (U251, HeLa, MCF-7). In addition, Thamnarak et al. [33] isolated two new ortholignans siamensinols (1–2) and seven known compounds agatharesinol (3), syringaresinol-glucoside (4), noreugenin from Selaginella siamensis (5), 8- methyleugenitol (6), melachromone (7), uncinoside A (8), and daucosterol (9). Among them, compounds 1–2 showed moderate inhibitory effects on MOLT-3 cells, while compounds 6-8 showed moderate inhibitory effects on three tumor cells (HepG2, A549 and HuCCA-1).

Biflavonoids are the primary active ingredients of Selaginella plants, exerting anti-cancer effects [26]. SFB species exert anti-tumor effects by inhibiting cancer cell proliferation, inducing cancer cell apoptosis, inhibiting tumor metastasis and angiogenesis, inducing autophagy, and impacting the tumor microenvironment. As shown in Figure 2. SFB has been shown to have anti-cancer effects on a variety of tumors, including ovarian cancer, lung cancer, prostate cancer, breast cancer and digestive system tumors.

Biflavonoids, a unique class of flavonoids, are primarily soluble in ethyl acetate. Their basic structural unit is made up of two molecules of flavonoids, generally in accordance with a C–C or C–O–C link. Currently, 71 different types of biflavonoids have been discovered in Selaginella plants [34]. They can be divided into five categories based on the various connection methods. As shown in Figure 3: ① amentoflavone-type (C′3-C″8)(I), of which there are 24 species, mainly represented by amentoflavone and isoginkgetin; ② robustaflavone-type (C′3-C″6)(II), with 23 species, mainly represented by robustaflavone; ③ hinokiflavone-t (C′4-O-C″6)(III), with ten species, mainly represented by hinokiflavone and isocryptomerin; ④ delicaflavone-type (C3-O-C4‴)(IV), of which there are four species, mainly represented by delicaflavone; and ⑤ biphenylether-type (C′3-O-C‴4)(V), of which there are two species.

Over the past 5 years, scientists have discovered numerous novel biflavonoids in Selaginella plants [35]. Nine species of bioflavonoids from Selaginella doederleinii have been recently discovered [29,30,31]. As shown in Figure 4. Among these, compound 2 [36] showed modest selectivity to A549 cells and MCF-7 cells; compound 4 [37] demonstrated potent cytotoxicity against the human cancer cell lines SMMC-7721, A549, and MCF-7; and compounds 6–9 [38] considerably inhibited the proliferation of non-small cell lung cancer. The most active compound, 9, caused apoptosis and cycle arrest in A549 cells. A new robustaflavone, (2S, 2′S)-2,3,2′,3′-tetrahydrorobustaflavone 7,4′,4‴-trimethyl ether (10), was isolated from a 75% ethanolic aqueous extract of Selaginella uncinate [39]. Selaginella trichoclada was extracted with 70% ethanol, and five novel biflavonoids were discovered. Among them, compounds 11 and 12 [40] demonstrated moderate cytotoxicity to human cancer cell lines A549 and HepG2; compound 13 [41] was effective against breast cancer via the mitochondrial pathway; compound 14 [42] demonstrated moderate cytotoxicity to human cancer cell lines DU145, MCF-7, and PC3; and compound 15 [43] showed cytotoxicity to the human breast cancer cell line MCF-7. A new biflavonoid (16) was isolated from a 70% ethanolic extract of Selaginella braunii, which showed significant antiproliferative effects on SMMC-7721, MCF-7, and A549 cells [44]. From Selaginella siamensis and Selaginella bryopteris, three novel biflavonoids known as siamamenflavones A–C (17–19) were isolated. One of them, siamamenflavone B (18), exhibited potent inhibition of wild-type EGFR protein [45]. Selaginella tamariscina yielded three novel biflavonoids: involvenflavone G (20), H (21), and I (22); H and I may have potent hypoglycemic effects [46].

Biflavonoids derived from plants other than those of the genus Selaginella also have significant anti-tumor effects. According to reports, inhibiting Eg5 can cause apoptosis and mitotic arrest. Eg5 inhibitors are therefore becoming the focus of anticancer drug research. As a mitotic kinesin Eg5 inhibitor, morelloflavone from Garcinia dulcis binds to the variable site of Eg5, thereby inhibiting the ATPase activity and motor function of Eg5 [47]. Chamaejasmenin isolated from Stellera chamaejasme L. roots inhibited the proliferation of cells from eight human solid tumor cell lines (HepG2, SMMC-7721, A549, MG63, U-2 OS, and KHOS, HCT-116 and HeLa) [48]. Japoflavone D, a biflavonoid from Lonicera japonica flower buds with significant antioxidant activity, has a dual regulatory effect on the apoptosis of liver cancer cells under various oxidative circumstances [49]